Enzymes 185 



fications were published by Manheimer and Seligman,^^ by 

 Loveless and Danielli,^^ and by Gomori.^^ The first two 

 methods use /3-naphthyl phosphate as a substrate; Loveless 

 and Danielli, a complicated azo dye phosphate; Gomori, a- 

 naphthyl phosphate. The method of Gomori is the simplest; 

 it employs only commerically available chemicals and does 

 not require cooling by ice. The pictures obtained with it are 

 remarkably sharp and much more detailed than those given 

 by methods using ^-naphthyl phosphate. 



Method 



Fixation and embedding as in the Ca-Co method. 



Protection with collodion is better avoided because the 

 membrane may be stained rather intensely by decomposi- 

 tion products. 



Composition of the incubating mixture.— Dissolve about 10 

 mg. of Na a-naphthyl phosphate ( available from Dajac Lab- 

 oratories, 511 Lancaster Ave., Leominster, Mass.) in a few 

 ml. of distilled water. Add a few ml. of a 4-5 per cent solu- 

 tion of borax (Na2B4O7*10H2O), about 40 ml. of cool dis- 

 tilled water (not warmer than 20° C.), and a few drops of a 

 10 per cent solution of magensium chloride or sulfate. Stir 

 into the mixture 20-50 mg. of any of the following diazonium 

 salts: Blue B, Red RC, Bordeaux GP, Red G, or Fast Blue 

 RR. 



Incubate the slides for 10 to 30 minutes or until the desired 

 intensity of staining is obtained. The slides may be removed 

 from the incubating mixture and inspected under the micro- 

 scope repeatedly. Mechanical stirring of the solution during 

 incubation is highly advisable ( p. 172 ) . 



Wash slides; counters tain with hematoxylin or with alum 

 carmine; diflFerentiate with acid alcohol (alcohol concentra- 

 tion, 70-80 per cent ) ; wash again and either mount in glyc- 

 erol-jelly or dehydrate and mount in balsam (p. 170). Sites 



50. Manheimer, L. H., and Seligman, A. M.: J. Nat. Cancer Inst., 9:181, 

 1948. 



51. Loveless, A., and Danielli, J. F.: Quart. J. Micr. Sc., 90:57, 1949. 



